Assembly of basement membrane in vitro by cooperation between alveolar epithelial cells and pulmonary fibroblasts

Cell Struct Funct. 1997 Dec;22(6):603-14. doi: 10.1247/csf.22.603.

Abstract

To investigate basement membrane formation by cooperation between pneumocytes and pulmonary fibroblasts, we cultured type II alveolar epithelial cells obtained from rats transfected with SV40-large T antigen gene (SV40-T2 cells) on type I collagen matrices. On fibroblasts-embedded gel (T2-Fgel), SV40-T2 cells ultrastructurally formed a continuous and thin layer of lamina densa, while on collagen gel without fibroblasts (T2-gel) SV40-T2 cells produced only discontinuous and diffuse deposits. Stripping SV40-T2 cells off the tissues by H2O2 treatment revealed a continuous and plane surface of lamina densa assembled on the T2-Fgel tissue, whereas only amorphous deposits appeared on the T2-gel tissue. Immunolocalization of major basement membrane components showed that type IV collagen, laminin, perlecan and entactin (nidogen) were continuously integrated on the lamina densa in T2-Fgel. In T2-gel, all these components were discontinuously distributed beneath SV40-T2 cells. The contribution of pulmonary fibroblasts to the assembly of basement membrane through reorganization of collagen matrix and/or soluble factors was examined by the cultured of SV40-T2 cells on the freeze-thawed fibroblast-tissue and/or with the fibroblast-conditioned medium. Both SV40-T2 cells on the freeze-thawed fibroblast-tissue and SV40-T2 cells in T2-gel in the fibroblast-conditioned medium failed to produce a lamina densa. SV40-T2 cells could assemble a lamina densa only on the freeze-thawed fibroblast-tissue in the fibroblast-conditioned medium. These results show that the basement membrane components are assembled to a lamina densa by combination of the reorganization of collagen matrix and the supply of soluble factors by pulmonary fibroblasts.

MeSH terms

  • Animals
  • Antigens, Polyomavirus Transforming / genetics
  • Basement Membrane / chemistry
  • Basement Membrane / metabolism
  • Basement Membrane / ultrastructure
  • Cell Communication / physiology*
  • Cells, Cultured
  • Collagen / analysis
  • Epithelial Cells / cytology*
  • Epithelial Cells / ultrastructure
  • Extracellular Matrix / chemistry
  • Extracellular Matrix / metabolism
  • Fibroblasts / cytology
  • Fibroblasts / ultrastructure
  • Fluorescent Antibody Technique
  • Laminin / analysis
  • Microscopy, Electron
  • Microscopy, Electron, Scanning
  • Pulmonary Alveoli / cytology*
  • Rats
  • Transfection

Substances

  • Antigens, Polyomavirus Transforming
  • Laminin
  • Collagen